p63 is a member of the p53 gene family that is essential for the development of ectodermally derived structures whose morphogenesis is dependent on ectodermal–mesenchymal interactions. Stratified epithelia fail to differentiate and hair follicles, teeth, sweat glands, and mammary primordia are not formed in mice lacking p63 (Mills et al., 1999; Yang et al., 1999). Although p63-deficient models have been extremely informative for assessing the in vivo role of p63 and for illuminating p63 as a candidate in several human developmental syndromes (Celli et al., 1999; Ianakiev et al., 2000; McGrath et al., 2001), the perinatal lethality of mice that are homozygous for p63 null alleles limits the utility of these models for examining the function of p63 at later stages. We have therefore generated a conditional p63 allele that has an essential region of the gene flanked by loxP sites and tested this allele by crossing it to a strain that expresses Cre during early embryogenesis. Cre excision is efficient and inactivates p63, leading to a phenotype that is indistinguishable from p63 nullizygous mice. The p63 floxed allele (p63floxN) was generated by embryonic stem (ES) cell technology using a replacement vector that exchanged the endogenous p63 gene with a modified version that had loxP sites flanking exons encoding the DNA binding domain (DBD)(Schmale and Bamberger 1997; Osada et al., 1998; Yang et al., 1998)(Fig. 1A). Recombinant clones were identified by Southern analysis and the functionality of the integrated loxP sites was assessed by introducing a Creexpressing plasmid into+/p63floxN ES cells. G418 sibselection was used to assess the presence or absence of the neomycin resistance cassette and Southern analysis using a probe specific for exon 5 was used to identify clones that contained exons encoding the DBD. Clones falling into each of the three possible classes of recombination products were obtained, indicating that all three loxP sites that had been introduced during the gene targeting step were functional. Mice carrying the p63floxN allele were generated by standard procedures.+/p63floxN mice were intercrossed to generate p63floxN/p63floxN progeny (Fig. 1B). Mice homozygous for the p63floxN allele were obtained at the expected frequency, were viable and fertile, and were phenotypically indistinguishable from wild-type siblings